Streptococcus pneumoniae is a major cause of morbidity and mortality worldwide. The pneumococcus is able to colonize and replicate in a number of sites in the human host including the nasopharynx (colonization), lungs (pneumoniae), blood (sepsis, and the brain (meningitis). Crucial to understanding the pathogenesis of pneumococcal disease is understanding the signals encountered by the bacteria and the specific transcriptional response to these signals. An unexplored setting is the high, potentially toxic, concentrations of calcium and other metals encountered as the pneumococcus enters the nasopharynx and translocates to the bloodstream. This application will elucidate the transcriptional response of S. pneumoniae to divalent cations by investigating the specific roles of environmental cations on global gene expression as well as characterize two newly discovered cation efflux systems that are required for host pathogenesis. In this study we will characterize the specific transcriptional responses of S. pneumoniae to calcium, manganese, and zinc by both microarray and qRT-PCR. This study provides a unique perspective to not only determine the global response to extracellular cation sensing, but since mutants in these transporters accumulate high levels of their cognate cation, allows for the determination of how intracellular cation concentration can influence cell signaling pathways. These data will be used to further understand the specific responses of S. pneumoniae to the various signals encountered in the host. The transporters themselves will be subject to extensive molecular characterization including determining conserved domains via sequencing clinical strain and then generating mutants based on sequence homology to ascertain critical residues for metal specificity and protein function. The knowledge gained by characterizing these transporters at a molecular level will aid in our understanding of metal transport in bacteria. PUBLIC HEALTH RELEVANCE: A majority of previous studies in pathogenic bacteria has focused on metal acquisition, in contrast our understanding of metal efflux by pathogenic bacteria remains an intriguing question. Furthermore, as these transporters are required for host pathogenesis, they provide a novel antibacterial target. By understanding the residues conferring metal ion selectivity and function, these studies will provide greater insight into the strategies utilized by pathogenic bacteria to export cations found in abundance in the human host.